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Topic:Biotechnology
Biotechnology in horses encompasses the application of biological techniques and tools to enhance equine health, performance, and reproduction. This field includes genetic engineering, cloning, and the development of vaccines and therapeutics tailored to equine physiology. Techniques such as gene editing and stem cell therapy are explored for their potential to address genetic disorders, improve tissue regeneration, and enhance disease resistance in horses. Additionally, advancements in reproductive biotechnology, such as artificial insemination and embryo transfer, contribute to genetic diversity and breeding efficiency. This page compiles peer-reviewed research studies and scholarly articles that investigate the applications, methodologies, and implications of biotechnology in equine science.
Equine embryos and embryonic stem cells: defining reliable markers of pluripotency. Cartilage and tendon injuries are a significant source of animal wastage and financial loss within the horse-racing industry. Moreover, both cartilage and tendon have limited intrinsic capacity for self-repair, and the functionally inferior tissue produced within a lesion may reduce performance and increase the risk of reinjury. Stem cells offer tremendous potential for accelerating and improving tissue healing, and adult mesenchymal stem cells (MSCs) are already used to treat cartilage and tendon injuries in horses. However, MSCs are scarce in the bone marrow isolates used, have limited poten...
Detection of prohibited animal products in livestock feeds by single-strand conformation polymorphism analysis. Single-strand conformation polymorphism (SSCP) analysis of amplicons produced from a mitochondrial DNA region between the tRNA(Lys) and ATPase8 genes was applied for the detection of animal product within livestock feeds. Identification of prohibited animal (cattle, elk, sheep, deer, and goat) and nonprohibited animal (pig and horse) products from North America was possible based on the differential display of the single-stranded DNA fragments for the different animal species on SSCP gels. This method allowed specific detection and identification of mixed genomic DNA from different animal spec...
Evaluation of early cellular influences of bone morphogenetic proteins 12 and 2 on equine superficial digital flexor tenocytes and bone marrow-derived mesenchymal stem cells in vitro. To evaluate early cellular influences of bone morphogenetic protein (BMP)12 and BMP2 on equine superficial digital flexor tenocytes (SDFTNs) and equine bone marrow-derived mesenchymal stem cells (BMDMSCs). Methods: 9 adult clinically normal horses. Methods: BMDMSCs and SDFTNs were cultured in monolayer, either untreated or transduced with adenovirus encoding green fluorescent protein, adenovirus encoding BMP12, or adenovirus encoding BMP2. Cytomorphologic, cytochemical, immunocytochemical, and reverse transcriptase-quantitative PCR (RT-qPCR) analyses were performed on days 3 and 6. Genetic pro...
Automated counting of nucleated cells in equine synovial fluid without and with hyaluronidase pretreatment. Microscopy is usually used to obtain manual total and differential cell counts in equine synovial fluid. A faster, more precise method is desirable. Objective: The objectives were to compare an automated impedance method with a manual method for obtaining total and differential cell counts in equine synovial fluid and to evaluate the effect of pretreatment with hyaluronidase on automated results. Methods: Synovial fluid samples (n=48) were collected into EDTA and analyzed within 48 hours. Automated total and differential cell counts were evaluated using a Medonic CA620-VET hematology analyzer ...
Tissue engineering in wound repair: the three “R”s–repair, replace, regenerate. Horses are predisposed to traumatic wounds that can be labor intensive and expensive to manage. Skin has a considerable potential for efficient and functional repair however, while cutaneous repair is a regenerative process in the fetus, this capability declines in late gestation as inflammation and scarring alter the outcome of healing. The historical gold standard for replacement of lost skin is the autologous skin graft. However, the horse's lack of redundant donor skin limits the practicality of full-thickness grafting to smaller wounds; moreover, graft failure is relatively common in equi...
Comparison of direct and indirect methods of intra-abdominal pressure measurement in normal horses. To develop a direct method for measuring intra-abdominal pressures in the standing horse, identify a reference interval for direct intra-abdominal pressures, compare these pressures to indirect intra-abdominal pressures measured from the bladder, and determine the optimal bladder infusion volume for indirect pressure measurement. Methods: Prospective, experimental study. Methods: A university-based equine research facility. Methods: Ten healthy adult horses, 5 males and 5 females. Methods: Direct intra-abdominal pressures were measured through an intraperitoneal cannula and zeroed at the heigh...
Evaluation of shape variability of stallion sperm heads by means of image analysis and Fourier descriptors. This study quantified and evaluated the variability of sperm head shape for 10 different stallions. Sperm head shape characteristics including sperm head length to width ratio, position of the center of gravity, curvature, and degree of roundness were assessed and analysed from images using elliptic Fourier descriptors and inverse Fourier transformation. The first four principal components accounted for 88.46-92.33% of the total variance and provided a good summary of the overall data. In the case of the ejaculate with defective sperm heads the components accounted for 97.35-98.21% of variatio...
Identification of horse chestnut coat color genotype using SNaPshot. The Cantabrian Coast horse breeds of the Iberian Peninsula have mainly black or bay colored coats, but alleles responsible for a chestnut coat color run in these breeds and occasionally, chestnut horses are born. Chestnut coat color is caused by two recessive alleles, e and e(a), of the melanocortin-1 receptor gene, whereas the presence of the dominant, wild-type E allele produces black or bay coat horses. Because black or bay colored coats are considered as the purebred phenotype for most of the breeds from this region, it is important to have a fast and reliable method to detect alleles caus...
Development of a blocking ELISA using a recombinant glycoprotein for the detection of antibodies to vesicular stomatitis New Jersey virus. A recombinant glycoprotein (R-GP) of vesicular stomatitis New Jersey virus (VSV-NJ) was expressed in insect cells by a baculovirus system. Its utility as a diagnostic antigen in a blocking ELISA was investigated as an alternative to the current native GP extracted from VSV-NJ. With the cut-off value of 73% inhibition, the R-GP ELISA exhibited 99.1% specificity for naive sera from cattle and horses. It did not cross-react with VSV-Indiana (VSV-IN) positive sera and differentiated from foot-and-mouth disease and swine vesicular disease. Taken together, this is the first report that the R-GP has ...
Selection of reference genes for quantitative real-time PCR in equine in vivo and fresh and frozen-thawed in vitro blastocysts. Application of reverse transcription quantitative real-time polymerase chain reaction is very well suited to reveal differences in gene expression between in vivo and in vitro produced embryos. Ultimately, this may lead to optimized equine assisted reproductive techniques. However, for a correct interpretation of the real-time PCR results, all data must be normalized, which is most reliably achieved by calculating the geometric mean of the most stable reference genes. In this study a set of reliable reference genes was identified for equine in vivo and fresh and frozen-thawed in vitro embryos....
Identification of variables that optimize isolation and culture of multipotent mesenchymal stem cells from equine umbilical-cord blood. OBJECTIVE-To optimize the isolation and culture of mesenchymal stem cells (MSCs) from umbilical-cord blood (UCB), identify variables that predicted successful MSC isolation, and determine whether shipping, processing, and cryopreservation altered MSC viability, recovery rates, and expansion kinetics. SAMPLE POPULATION-UCB samples from 79 Thoroughbred and Quarter Horse mares. PROCEDURES-UCB samples were processed to reduce volume and remove RBCs. Nucleated cells (NCs) were cryopreserved or grown in various culture conditions to optimize MSC monolayer expansion and proliferation. Donor and UCB-s...
Detection of treponemes in canker lesions of horses by 16S rRNA clonal sequencing analysis. Equine canker is a chronic pododermatitis of the hoof in horses. Although spirochetes are detectable histopathologically in the lesions, the precise etiology remains unclear. This study reports the 16S rRNA gene sequencing of randomly selected clones based on PCR with Treponema-specific primers, using the canker lesions from two horses and healthy frog and sole from a horse. A total of 114 clones were obtained from the lesions, but no clones were detected in the healthy hoof tissues. The clones from the canker lesions examined were grouped into 19 operational taxonomic units, such as treponema...
Development of a novel equine whole transcript oligonucleotide GeneChip microarray and its use in gene expression profiling of normal articular-epiphyseal cartilage. No large scale equine microarray is available commercially to allow genomic and transcriptional profiling of the majority of genes that would define the genetic basis of equine disease. Objective: To generate a whole transcript target labelled GeneChip to interrogate the equine transcriptome and validate chip performance using RNA samples derived from organs, articular cells and normal cartilage. Methods: Equine mRNA and selected equine gene sequences derived from perfect cross-hybridisation of equine RNA on human microarray GeneChips, were used to design a custom equine gene microarray. Seque...
Establishing a reproducible method for the culture of primary equine corneal cells. To establish a reproducible method for the culture of primary equine corneal epithelial cells, keratocytes, and endothelial cells and to describe each cell's morphologic characteristics, immunocytochemical staining properties and conditions required for cryopreservation. Methods: Corneas from eight horses recently euthanized for reasons unrelated to this study were collected aseptically and enzymatically separated into three individual layers for cell isolation. The cells were plated, grown in culture, and continued for several passages. Each cell type was characterized by morphology and immun...
Computerised analysis of standardised ultrasonographic images to monitor the repair of surgically created core lesions in equine superficial digital flexor tendons following treatment with intratendinous platelet rich plasma or placebo. The effectiveness of new therapies to treat tendon injuries is difficult to determine and is often based on semi-quantitative methods, such as grey level analysis of ultrasonographic images or subjective pain scores. The alternatives are costly and long-lasting end-stage studies using experimental animals. In this study, a method of ultrasonographic tissue characterisation (UTC), using mathematical analysis of contiguous transverse ultrasonographic images, was used for intra-vital monitoring of the healing trajectory of standardised tendon lesions treated with platelet rich plasma (PRP) or pla...
Identification of Western equine encephalitis virus structural proteins that confer protection after DNA vaccination. DNA vaccines encoding different portions of the structural proteins of western equine encephalitis virus were tested for the efficacy of their protection in a 100% lethal mouse model of the virus. The 6K-E1 structural protein encoded by the DNA vaccine conferred complete protection against challenge with the homologous strain and limited protection against challenge with a heterologous strain.
Probing the calcium and sodium local environment in bones and teeth using multinuclear solid state NMR and X-ray absorption spectroscopy. Despite the numerous studies of bone mineral, there are still many questions regarding the exact structure and composition of the mineral phase, and how the mineral crystals become organised with respect to each other and the collagen matrix. Bone mineral is commonly formulated as hydroxyapatite, albeit with numerous substitutions, and has previously been studied by (31)P and (1)H NMR, which has given considerable insight into the complexity of the mineral structure. However, to date, there has been no report of an NMR investigation of the other major component of bone mineral, calcium, nor of...
Cloning, sequencing and expression analysis of the equine hepcidin gene by real-time PCR. Equine serum or plasma iron concentration drops quickly during inflammation. Accumulation of iron inside macrophages and reduction of the intestinal absorption of this element cause hypoferremia during systemic inflammatory processes. These mechanisms are mediated by hepcidin, a 25 amino acids peptide synthesized mainly in the liver in response to iron stores and inflammation. Hepcidin is an important peptide for systemic iron homeostasis and also has antibacterial and antifungal activities. Hepcidin up-regulation is particularly useful during acute inflammation, especially before adaptive imm...
Horse breed discrimination using machine learning methods. Genetic relationships and population structure of 8 horse breeds in the Czech and Slovak Republics were investigated using classification methods for breed discrimination. To demonstrate genetic differences among these breeds, we used genetic information - genotype data of microsatellite markers and classification algorithms - to perform a probabilistic prediction of an individual's breed. In total, 932 unrelated animals were genotyped for 17 microsatellite markers recommended by the ISAG for parentage testing (AHT4, AHT5, ASB2, HMS3, HMS6, HMS7, HTG4, HTG10, VHL20, HTG6, HMS2, HTG7, ASB17, AS...
Abdominal wall reconstruction using biological tissue grafts. Synthetic mesh products have been used to repair abdominal wall defects (eg, hernias) for many years. Biological mesh products are now available as an option when synthetic mesh products are not appropriate. To correctly prepare biological tissue grafts for use in the OR, perioperative nurses must understand the types of grafts available. Biological tissue grafts may be harvested from human, porcine, bovine, or equine hosts and from skin, pericardium, or small intestine submucosa.
Evaluation of prolactin receptor (PRLR) as candidate gene for male fertility in Hanoverian warmblood horses. Stallion fertility has increasing importance as the artificial insemination is employed in horses more intensely. Molecular genetic markers may be useful tools to evaluate the stallion fertility before breeding. The prolactin receptor gene (PRLR) was chosen as a candidate for stallion fertility because of its influence on testicular and accessory sex gland function. Screening the equine PRLR gene for polymorphisms in Hanoverian stallions revealed two single nucleotide polymorphisms (SNPs). Association and haplotype analyses were performed in 162 Hanoverian warmblood stallions for these intrage...
Zonal chondrocyte subpopulations reacquire zone-specific characteristics during in vitro redifferentiation. If chondrocytes from the superficial, middle, and deep zones of articular cartilage could maintain or regain their characteristic properties during in vitro culture, it would be feasible to create constructs comprising these distinctive zones. Objective: Zone-specific characteristics of zonal cell populations will disappear during 2-dimensional expansion but will reappear after 3-dimensional redifferentiation, independent of the culture technique used (alginate beads versus pellet culture). Methods: Controlled laboratory study. Methods: Equine articular chondrocytes from the 3 zones were expan...
Cryopreservation does not affect the stem characteristics of multipotent cells isolated from equine peripheral blood. Mammalian adult stem cells show, in vitro, extensive differentiative ability and may represent a versatile tool for tissue regenerative purposes, even after long-term storage. Multipotent stem cells isolated from horse blood have been shown to possess the capacity to differentiate into diverse mesenchymal lineages although their full characterization is still at an early stage. The aim of this study was to examine the effects of cryopreservation on stemness characteristics of adult equine mesenchymal stem cells isolated from peripheral blood (ePB-MSC). Each sample of ePB-MSC was analyzed immed...
Surface modified polymeric nanoparticles for immunisation against equine strangles. The successful development of particulate vaccines depends on the understanding of their physicochemical and biological characteristics. Therefore, the main purpose of this study was to develop and characterise stable surface modified poly(lactic acid) (PLA) nanoparticles, using polyvinyl alcohol (PVA), alginate (ALG) and glycolchitosan (GCS) containing a Streptococcus equi enzymatic extract adsorbed onto the surface. The characterisation of the preparations and a physicochemical study of the adsorption process were performed. The adsorption of S. equi proteins is a rapid process reaching, wit...
Structural and functional-annotation of an equine whole genome oligoarray. The horse genome is sequenced, allowing equine researchers to use high-throughput functional genomics platforms such as microarrays; next-generation sequencing for gene expression and proteomics. However, for researchers to derive value from these functional genomics datasets, they must be able to model this data in biologically relevant ways; to do so requires that the equine genome be more fully annotated. There are two interrelated types of genomic annotation: structural and functional. Structural annotation is delineating and demarcating the genomic elements (such as genes, promoters, and ...
The elastin network: its relationship with collagen and cells in articular cartilage as visualized by multiphoton microscopy. A combination of two-photon fluorescence (TPF), second harmonic generation (SHG) and coherent anti-Stokes Raman scattering (CARS) imaging has been used to investigate the elastin fibre network in healthy equine articular cartilage from the metacarpophalangeal joint. The elastin fibres were identified using their intrinsic two-photon fluorescence and immuno-staining was used to confirm the identity of these fibres. SHG was used to reveal the collagen matrix and the collagen fibre orientations were determined from their SHG polarization sensitivity, while CARS was used to clearly delineate the c...
Equine transcriptome quantification using human GeneChip arrays can be improved using genomic DNA hybridisation and probe selection. Affymetrix GeneChip arrays are a powerful tool for transcriptome profiling and have been applied to a wide range of species. A genomic DNA (gDNA)-based probe selection method has been developed which broadens the range of species to which GeneChips may be successfully applied. This study demonstrated that gDNA-based probe selection on the Affymetrix U133+2 GeneChip array can be used to study the equine transcriptome which, to date, has received only limited attention. More than 29,000 transcripts can be detected in equine brain and liver and in primary cultures of equine articular chondrocytes...
Dermal fibroblast-mediated BMP2 therapy to accelerate bone healing in an equine osteotomy model. This study evaluated healing of equine metacarpal/metatarsal osteotomies in response to percutaneous injection of autologous dermal fibroblasts (DFbs) genetically engineered to secrete bone morphogenetic protein-2 (BMP2) or demonstrate green fluorescent protein (GFP) gene expression administered 14 days after surgery. Radiographic assessment of bone formation indicated greater and earlier healing of bone defects treated with DFb with BMP2 gene augmentation. Quantitative computed tomography and biomechanical testing revealed greater mineralized callus and torsional strength of DFb-BMP2-treated ...
Cloning of IgE-binding proteins from Simulium vittatum and their potential significance as allergens for equine insect bite hypersensitivity. Insect bite hypersensitivity (IBH) is an allergic dermatitis of horses caused by bites of Culicoides and sometimes Simulium spp. The aim of this investigation was to identify Simulium allergens associated with IBH. A phage surface display cDNA library expressing recombinant Simulium vittatum salivary gland proteins was screened using sera of IBH-affected horses sensitized to S. vittatum salivary gland proteins as shown in immunoblot, resulting in the identification of seven cDNAs encoding IgE-binding proteins. The deduced amino acid sequences of these proteins showed sequence similarities to a...
